Project Summary - Project 1 Long-term success in the treatment of tobacco-related head and neck squamous cell carcinoma (HNSCC) is hindered by an alarming rate of second primary tumor (SPT) development following curative treatment. Patients with human papillomavirus (HPV)-negative HNSCC develop a SPT of the upper aerodigestive tract at the rate of 3-6% per year, and are most likely to succumb to these secondary cancers. Although smoking cessation reduces the occurrence of SPTs, moderation of risk is not observed for 5 years, and is insufficient to return risk to baseline. The availability of a well-tolerated and affordable intervention that prevents SPTs would have a major global impact on mortality and quality of life in patients at risk. Unfortunately, no tolerable and effective chemopreventive agents have been identified for HNSCC. Our broad, long-term goal is the rigorous translational development of a tolerable and effective chemoprevention strategy against HNSCC SPTs. Reduced risk for HNSCC and SPTs is associated with diets rich in the Brassica family of cruciferous vegetables, including broccoli. Broccoli is rich in glucoraphanin, which is metabolized to the key bioactive component sulforaphane (SF). SF induces the expression of the transcription factor NRF2, which leads to upregulation of NRF2 target genes. A number of NRF2 target genes encode cytoprotective enzymes, which act to detoxify environmental carcinogens including benzene, aldehydes and nitrosamines found in tobacco smoke. The relevance of the NRF2 signaling pathway for oral cancer chemoprevention is highlighted by the enhanced susceptibility of mice lacking the Nrf2 gene to oral cancer induced by the carcinogen 4NQO. We are developing broccoli seed preparations (BSPs) as a chemopreventive agent against carcinogen-induced cancers, and have determined the safety, tolerability, and pharmacokinetics of BSPs in humans. We have also shown that SF induces NRF2 and NRF2 target gene expression in normal oral keratinocytes and in HNSCC cell lines. Moreover, we have provided first-time demonstration that transcripts for NRF2 target genes are upregulated in the oral mucosa of healthy volunteers treated with SF-rich BSP. We hypothesize that NRF2 pathway activation in oral epithelium can be induced by administering BSP to patients curatively treated for a first tobacco-related HNSCC, and that the target level of NRF2 pathway activation for chemopreventive efficacy in humans can be determined in a mouse model of carcinogen-induced HNSCC. To test this hypothesis we propose two Specific Aims: 1) To investigate the dose-response relationship between sulforaphane (SF) and chemopreventive efficacy in a mouse model of carcinogen-induced HNSCC, and 2) To systematically assess the clinical chemopreventive potential of BSP administration to patients with tobacco- related HNSCC at high risk for SPT.